ABSTRACT The expanding development of image-guided minimally invasive thermal therapies could greatly benefit from improved quantitative imaging techniques. Although, non-invasive and non-ionizing magnetic resonance guided focused ultrasound (MRgFUS) treatments are commonly assessed with qualitative MRI measures, additional quantitative metrics have been investigated to provide more consistent, repeatable and accurate assessment of treatment outcomes. Sophisticated acquisition schemes and reconstruction methods now allow for the real-time, simultaneous mapping of several quantitative tissue parameters during MRgFUS treatments. However, none of these potentially promising metrics have been rigorously correlated to the state of tissue- specific response assessed at the cellular level with histopathology. Rigorous comparison with histology is critical, particularly when treating malignant disease. Unfortunately, orientation loss, deformation, and shrinkage of excised tissue impede voxel-based correlation of in vivo MRI to histological outcomes. As a result, the complex relationship between quantitative parameters maps from MRI and the final state of tissue is not yet known. This proposal seeks to address this issue through the development and preliminary validation of a multi-step MRI-to-histology registration methodology that creates an accurate mapping between the MRI image coordinates and the high-resolution data from histology. Although explicitly developed for MRgFUS treatments the method should be broadly applicable. The methodology will include rigid and non-rigid registration of: 1) histology slides to their block face images for the creation of a 3D histological model, 2) the 3D histological model to an ex vivo MRI model generated following surgical excision, and 3) the ex vivo MRI model to the in vivo MRI model acquired post MRgFUS ablation. This methodology will provide a critical tool not only for the development and translation of new MRgFUS treatments but for other interventional techniques that require correlation between MRI treatment assessment metrics and numerous gold standard histological stains.